CN112999412A

CN112999412A - Hydrogel dressing for wound healing and preparation method thereof

Info

Publication number: CN112999412A
Application number: CN202110260621.3A
Authority: CN
Inventors: 胡佳颖; 陈倩; 刘庄
Original assignee: Suzhou University
Current assignee: Huijiawang Tianjin Technology Co ltd; JIANGSU OPERA MEDICAL SUPPLIES CO Ltd
Priority date: 2021-03-10
Filing date: 2021-03-10
Publication date: 2021-06-22
Anticipated expiration: 2041-03-10
Also published as: CN112999412B

Abstract

The invention discloses a hydrogel dressing which is a semi-interpenetrating network hydrogel obtained by carrying out free radical polymerization on poly (N-isopropylacrylamide), polymethacrylic acid and gelatin. The invention also discloses a preparation method of the hydrogel dressing. The hydrogel dressing of the invention is composed of thermally responsive tough adhesive hydrogel, combines the characteristics of high ductility, toughness, tissue adhesion, macrophage reaction change and the like, and has the effects of mechanical activity and immunoregulation. The dressing adheres strongly to the skin and responds to exposure to skin temperature to actively contract the wound to accelerate wound healing. The new mechanical treatment and biological-free method is expected to be widely used in various medical environments.

Description

Hydrogel dressing for wound healing and preparation method thereof

Technical Field

The invention relates to the technical field of biological materials, in particular to a hydrogel dressing for wound healing and a preparation method thereof.

Background

Various types of wounds resulting from diabetes or circulatory disorders, including acute post-operative wounds, traumatic wounds, burns and chronic wounds, remain a major clinical concern and have significant impact on patients and society. Wound healing is an important physiological process involving evolutionarily conserved sequences of hemostasis, inflammation, proliferation and tissue remodeling. A key process during the wound healing response is the restoration of the epithelial layer to restore the integrity of the skin barrier. Although various wound dressings such as gauze, cotton linters and hydrogel have been widely used, existing adhesive wound dressings are not satisfactory for treating large area or chronic wounds due to their slow and passive healing behavior. To promote wound healing, efforts have been made to provide antimicrobial agents to protect wounds from pathogen infection, and to use bioactive agents (e.g., growth factors or cells) to accelerate cell proliferation and tissue remodeling. Nevertheless, such functional wound dressings are often subject to complicated manufacturing processes, potential side effects associated with the drug, high cost, handling and storage difficulties, and problems with encapsulated bioactive agent payloads and controlled release.

In addition to wound dressings that encapsulate antibacterial and bioactive agents to promote wound healing, modulation of innate immune cells, particularly macrophages, has also attracted a wide range of attention, the latter playing a key role in promoting tissue regeneration and tissue remodeling. Methacrylic acid (MAA) based materials are reported to polarize macrophages towards a regeneration-promoting (M2) phenotype. This polarization of macrophages can further regulate the secretion of inflammation-associated cytokines, such as interleukin-1 β (IL-1 β), interleukin-10 (IL-10), interleukin-6 (IL-6), and tumor necrosis factor- α (TNF- α), to promote tissue regeneration. In addition, MAA can promote wound healing by promoting neovascularization of surrounding tissues and modulating angiogenesis-related factors, including platelet-derived growth factor-B (PDGF-B), basic fibroblast growth factor (bFGF), thrombospondin-1 (TSP-1), Vascular Endothelial Growth Factor (VEGF), and CXC motif chemokine-10 (CXCL-10). Thus, designing an immunomodulatory wound dressing may be an interesting strategy for promoting wound healing without the need for additional drugs.

Adhesive hydrogels having long-term adhesion, excellent mechanical properties and high biocompatibility have attracted considerable attention as wound dressings. However, conventional wound dressings that passively promote healing often fail to exhibit desirable efficacy. Although various drugs or cells have been introduced to accelerate the healing process, this strategy is often limited, including side effects of the drugs, complicated preparation processes, and high costs.

Disclosure of Invention

The technical problem to be solved by the invention is to provide a hydrogel dressing which is composed of thermal response tough adhesive hydrogel, combines the characteristics of high ductility, toughness, tissue adhesion, macrophage reaction change and the like, and has the effects of mechanical activity and immunoregulation. The dressing adheres strongly to the skin and responds to exposure to skin temperature to actively contract the wound to accelerate wound healing. The new mechanical treatment and biological-free method is expected to be widely used in various medical environments.

In order to solve the technical problems, the invention provides the following technical scheme:

the invention provides a hydrogel dressing which is semi-interpenetrating network hydrogel (PNIPAm-P) obtained by performing free radical polymerization reaction on poly (N-isopropylacrylamide), polymethacrylic acid and gelatin.

Poly (N-isopropylacrylamide) (Poly (N-isoproyl Acrylamide), PNIPAm for short) is polymerized from the monomer N-isopropylacrylamide (NIPAm). The phase change occurs when the temperature of the PNIPAm aqueous solution is raised to about 32 ℃, the homogeneous system is changed into the heterogeneous system, and the volume of the chemically crosslinked PNIPAm hydrogel suddenly shrinks when the temperature is raised to about 32 ℃. When the macromolecular chain of PNIPAm has hydrophilic amido and hydrophobic isopropyl, the water solution of linear PNIPAm and the cross-linked PNIPAm hydrogel have temperature sensitive characteristic, so that the heat sensitive polymer material is widely applied in the aspects of drug controlled release, biochemical separation, chemical sensor, etc.

Methacrylic Acid (MAA) based materials are angiogenic materials that promote wound healing. They can be made in different forms, such as beads, porous scaffolds, hydrogels and lubricious coatings. More importantly, MAA-based materials do not contain bioactive agents, but they have biological effects. Recent studies have elucidated the mechanism of action of MAA. Macrophages that interact with this biomaterial are predisposed to the anti-inflammatory phenotype (M2) and increase insulin growth factor-1 (IGF-1) secretion. This secretion increases the proliferation and migration of endothelial cells, consistent with higher vascular density and improved healing in the surrounding tissue. Furthermore, MAA-based materials do not exhibit significant foreign body reactions. Thus, such materials exhibit a wide range of biomedical applications.

Gelatin (Gelatin, Gel for short), a macromolecular hydrocolloid, is the product of partial hydrolysis of collagen. Colorless to light yellow solid, and the relative molecular mass is about 50000-100000. The relative density is 1.3 to 1.4. Is insoluble in water, but when soaked in water, it can absorb 5-10 times of water to swell and soften, and when heated, it dissolves into colloid, and when cooled to 35-40 deg.C, it becomes gel. Because of high biocompatibility and biodegradability, no other byproducts are generated after in vivo degradation, no immunogenicity and blood compatibility, and the same components and biological properties as collagen, the collagen-based collagen is widely applied to tissue engineering and drug delivery systems.

The hydrogel dressing consists of poly (N-isopropylacrylamide), polymethacrylic acid and gelatin, and the semi-interpenetrating network hydrogel PNIPAm-P is obtained through free radical polymerization. The hydrogel strongly adheres to the skin through an amino/carboxyl reaction and then actively contracts the wound due to the responsiveness of the component PNIPAm in the hydrogel to skin temperature. PMAAs are inserted as long polymer chains in the PNIPAm hydrogel network, promote wound healing and neovascularization by endogenous mechanisms including macrophage polarization. Gelatin is used as a natural polymer material, so that the PNIPAm-P hydrogel has better ductility, toughness, tissue adhesion and biocompatibility. In vitro and in vivo studies have shown that they are very effective in accelerating and supporting the healing of skin wounds. The mechano-active and immunomodulatory adhesive dressing PNIPAm-P of the invention is therefore an ideal medical product.

Further, in the raw materials for synthesizing the hydrogel dressing, the mass ratio of poly (N-isopropylacrylamide), polymethacrylic acid and gelatin is 1: 1: 1-4: 1: 1, preferably 4: 1: 1.

further, at 25 ℃, the pore diameter of the hydrogel dressing is 7-8 μm; the pore diameter of the hydrogel dressing is 4-5 mu m at 37 ℃. The significant reduction in pore size indicates that the PNIPAm-P containing hydrogels have excellent temperature sensitive shrink properties. Preferably, the aperture of the PNIPAm-P hydrogel is 7-8 μm.

In a second aspect, the present invention provides a method for preparing a hydrogel dressing according to the first aspect, comprising the steps of:

adding sodium polymethacrylate, cross-linking agent and gelatin solution into poly (N-isopropylacrylamide), mixing, and introducing N₂Deoxidizing, then adding an initiator and an accelerator, sealing, and carrying out polymerization reaction for 6-8h at the temperature of 0-40 ℃ to obtain the hydrogel dressing.

In the invention, when the PNIPAm-P hydrogel is prepared, the PNIPAm-P hydrogel with different masses of the NIPAm, the gelatin and the PMAA-Na can be obtained by controlling the amount of the gelatin and the PMAA-Na powder added into the aqueous solution of the NIPAm.

Furthermore, the molecular weight of the polymethacrylic acid is 6-100 kDa, and the concentration of the solution is 25 mg/mL.

Further, the concentration of the gelatin solution is 25mg/mL, and the concentration of the poly (N-isopropylacrylamide) solution is 100 mg/mL.

Further, the cross-linking agent is a common water-soluble cross-linking agent, such as N, N' -methylene bisacrylamide, ethylene glycol diacrylate, m-phenylene bismaleimide, pentaerythritol triacrylate, and the like.

Further, the initiator is a commonly used water-soluble initiator such as persulfate of inorganic salts, hydrogen peroxide, a water-soluble azo initiator, etc., and the accelerator is a commonly used accelerator for redox reaction such as tetramethylethylenediamine, triethylamine, etc.

Further, the solvents for dissolving poly (N-isopropylacrylamide), polymethacrylic acid and gelatin were all ultrapure water.

Further, after gelation, the method further comprises the steps of immersing the obtained hydrogel in saline water to remove impurities such as an excess initiator and an accelerator during the reaction, and then freeze-drying.

In use, the hydrogel dressing of the present invention can be used to seal a lyophilized hydrogel and a sterile saline solution in two separate chambers of a commercially available package for wound treatment in a ready-to-use form.

The invention has the beneficial effects that:

1. the invention is inspired by the contraction of chicken embryo wounds and utilizes the recent application in hydrogels and adhesives to design PNIPAm-P hydrogel dressings with mechanical activity and immunomodulatory properties. The PNIPAm-P hydrogel dressing consists of a thermal response polymer poly (N-isopropylacrylamide), polymethacrylic acid and gelatin, and PMAA which is inserted in a hydrogel network can further accelerate wound healing by promoting the formation of new blood vessels and regulating inflammatory reaction. The PNIPAm-P hydrogel dressing combines temperature-sensitive contraction and immunoregulation, and provides a new way for wound treatment.

2. The PNIPAm-P hydrogel dressing has a number of unique advantages over existing tissue dressings, including strong adhesion to tissue through covalent cross-linking, thermo-responsive contractile ability to mechanically contract wounds, modulation of inflammation, promotion of neovascularization, and ease of storage and use.

3. The PNIPAm-P can effectively accelerate the wound healing process of a full-thickness skin defect mouse model and a partial-thickness skin defect pig wound model through the following synergistic effect, including strong adhesion to tissues, mechanical wound contraction, angiogenesis promotion, collagen deposition and inflammation reduction.

4. The PNIPAm-P hydrogels of the present invention also exhibit a wide range of applications in other biomedical fields, such as diabetic wound healing and tissue defect repair. Thus, such hydrogel dressings, which are easy to prepare and easy to store, may offer new opportunities for wound healing and indicate new directions in the fields of bioscaffolds, drug delivery, and wearable or implantable medical devices.

5. The preparation method of the PNIPAm-P dressing is simple to operate, the raw materials are easy to obtain, and the PNIPAm-P dressing can be used for large-scale preparation of the PNIPAm-P dressing.

Drawings

FIG. 1 is a graph of the compressive properties of PNIPAm-P hydrogels;

FIG. 2 is a representative scanning electron microscope image of PNIPAm-P hydrogel at different temperatures;

FIG. 3 is a graph of the change in area and volume of a PNIPAm-P hydrogel at equilibrium conditions at 37 ℃;

FIG. 4 is a graph of the in vitro biocompatibility of PNIPAm-P hydrogel: (a) is a fluorescence light microscope micrograph of fibroblasts on PNIPAm-P hydrogel, (b) is an MTT method for analyzing the proliferation of fibroblasts;

FIG. 5 is a graph of wound contraction effect of PNIPAm-P hydrogel on fresh rodent skin tested in vitro;

fig. 6 is a graph of wound healing after dressing treatment: (a) representative images of wounds on

days

0, 2, 4 and 6 of treatment, (b) quantitative statistics of (a);

FIG. 7 is a graph of macrophage changes at the wound site: (a) and (b) is a flow cytometer analysis representative image and quantitative statistics of macrophages (CD11b + F4/80+), (c) and (d) is a flow cytometer analysis representative image and quantitative statistics of M2-class macrophages (CD206+ CD11b + F4/80 +);

FIG. 8 is a graph showing the expression of inflammation-associated cytokines on day 5 of wound treatment;

figure 9 is a graph of angiogenesis after dressing treatment: (a) is a representative photoacoustic image in neonatal wound tissue, (b) is a quantitative statistic of oxyhemoglobin saturation;

fig. 10 is a graph of wound healing after dressing treatment: (a) representative images of wounds at

day

0, 2, 4, 6 and 7 of treatment, (b) is the quantitative statistic of (a);

fig. 11 is a graph of wound healing after dressing treatment: (a) representative images of wounds on

days

0, 2, 4 and 6 of treatment, (b) quantitative statistics of (a);

FIG. 12 is a graph showing the effect of PNIPAm-P contraction on a defective region of an organ in vitro.

Detailed Description

The present invention is further described below in conjunction with specific examples to enable those skilled in the art to better understand the present invention and to practice it, but the examples are not intended to limit the present invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items.

Example 1: preparation of PNIPAm-P hydrogel

First, polymethacrylic acid (PMAA) (6kDa) was synthesized by conventional free radical polymerization and incubated overnight with NaOH to obtain PMAA-Na as sodium salt. Next, a NIPAm solution (solution A) was prepared at a concentration of 100mg/mL by dissolving an amount of N-isopropylacrylamide (NIPAm) in water. Then, a gelatin solution (solution B) having a concentration of 25mg/mL was prepared by dissolving an amount of gelatin in water. Next, solution B, PMAA-Na (25mg/mL) and crosslinker Bis were slowly added to solution A (solution C). After uniform mixing, the solution C is placed in N₂Bubbling down for 30 minutes to remove oxygen. Finally, initiator APS and accelerator TEMED were added to solution C and reacted at 4 ℃ for 6-8 hours to obtain PNIPAm-P hydrogel. After gelation, the hydrogel was removed and cut into small pieces, soaked in PBS for 30 minutes, and rinsed 3 times before use. The hydrogel can be lyophilized for storage.

Example 2: preparation of PNIPAm hydrogel

First, a NIPAm solution (solution a) was prepared at a concentration of 100mg/mL by dissolving an amount of N-isopropylacrylamide (NIPAm) in water. Next, a gelatin solution (solution B) having a concentration of 25mg/mL was prepared by dissolving an amount of gelatin in water. After the solutions A and B are mixed uniformly, in N₂Bubbling down for 30 minutes to remove oxygen. Finally, initiator APS and accelerator TEMED were added to the mixed solution and reacted at 4 ℃ for 6-8 hours to obtain PNIPAm hydrogel. After gelation, the hydrogel was removed and cut into small pieces, which were soaked in PBSSoak for 30 minutes, rinse 3 times before use. The hydrogel can be lyophilized for storage.

Example 3: preparation of PAAm-P hydrogel

First, polymethacrylic acid (PMAA) (6kDa) was synthesized by conventional free radical polymerization and incubated overnight with NaOH to obtain PMAA-Na as sodium salt. Next, an AAm solution (solution D) having a concentration of 100mg/mL was prepared by dissolving an amount of acrylamide (AAm) in water. Then, a gelatin solution (solution B) having a concentration of 25mg/mL was prepared by dissolving an amount of gelatin in water. Next, solution B, PMAA-Na (25mg/mL) and crosslinker Bis were slowly added to solution D (solution E). After mixing well, solution E is placed in N₂Bubbling down for 30 minutes to remove oxygen. Finally, initiator APS and accelerator TEMED were added to solution E and reacted at 4 ℃ for 6-8 hours to obtain PAAm-P hydrogel. After gelation, the hydrogel was removed and cut into small pieces, soaked in PBS for 30 minutes, and rinsed 3 times before use. The hydrogel can be lyophilized for storage.

Example 4: characterization of PNIPAm-P hydrogels

The compression properties of the hydrogels were tested by a TMS-Pro universal tester. The base area is 4.5cm²The cylindrical PNIPAm-P hydrogel was subjected to a compression test at a rate of 10mm/min, and the compression ratio was 50%. The morphological structure of these hydrogels was observed using scanning electron microscopy at 25 ℃ and 37 ℃. The thermo-response behavior of the hydrogel was evaluated by placing the hydrogel at 37 ℃ and measuring the change in volume over time. The initial and final dimensions are denoted as l_oAnd l, area strain from 1- (l)_o/l)²And (4) calculating. The results are shown in FIGS. 1-3.

FIG. 1 is a graph of the compressive properties of PNIPAm-P hydrogels. It is evident from the figure that the PNIPAm-P hydrogel can withstand high pressures to complete the deformation without rupture. The PNIPAm-P hydrogel has higher compressibility, which is probably due to the integration of gelatin in a PNIPAm network and the non-covalent interaction among the PNIPAm, the PMAA and the gelatin, dissipates energy under large deformation, and improves the mechanical property of the hydrogel.

FIG. 2 is a representative scanning electron microscope image of PNIPAm-P hydrogel at different temperatures. As can be seen from the figure, the pore size of the temperature sensitive hydrogel PNIPAm-P was significantly reduced at 37 deg.C compared to the hydrogel at 25 deg.C, indicating that the hydrogel containing PNIPAm had excellent temperature sensitive shrinkage properties.

FIG. 3 is a graph of the change in area and volume of a PNIPAm-P hydrogel at 37 ℃ under equilibrium conditions. As shown, the PNIPAm-P hydrogel shrunk to its initial volume in 40 minutes

Resulting in an area shrinkage of 90%. This unique temperature triggers the contractile behavior of this hydrogel, encouraging further exploration of its ability to mechanically contract the wound.

Example 5: toxicity study of PNIPAm-P hydrogel at cellular level

NIH-3T3 fibroblasts were cultured at 5X 10⁴Is seeded on a culture plate and cultured in an incubator for 2 hours to allow the cells to adhere. The hydrogel was purified by repeated soaking in PBS and 75% ethanol to remove excess TEMED and other residues prior to co-culturing with fibroblasts. Then, PNIPAm-P hydrogel was added to the wells, and 1mL of Dulbecco's modified Eagle's medium containing 10% fetal bovine serum was added to each well. Cells were allowed to adhere and grow for 1, 3 and 5 days. The morphology of the cells co-cultured with the hydrogel was observed using a fluorescence light microscope, and cell viability was evaluated by MTT analysis. The results are shown in FIG. 4.

FIG. 4 is a graph of the in vitro biocompatibility of PNIPAm-P hydrogel. FIG. 4(a) is a fluorescence light microscope micrograph of fibroblasts on PNIPAm-P hydrogel, and FIG. 4(b) is a micrograph of fibroblasts analyzed by MTT method for proliferation. The above results all indicate that PNIPAm-P hydrogel showed negligible effect on fibroblast proliferation.

Example 6: temperature-sensitive contraction research of PNIPAm-P hydrogel on in-vitro level

In vitro wound closure tests were performed on fresh explanted rodent skin (male BALB/c mice, 6-8 weeks). The PNIPAm-P hydrogel prepared in example 1 was applied to wound skin 8mm in diameter and lightly compressed at room temperature for 10 min. The skin and hydrogel were then left overnight at 37 ℃ and then rapidly frozen with liquid nitrogen to maintain wound size. Finally PNIPAm-P was removed and the wound was measured and analyzed by Image J software observation. The results are shown in FIG. 5.

FIG. 5 is a graph of wound contraction effect of PNIPAm-P hydrogel on fresh rodent skin tested in vitro. As shown, the PNIPAm-P hydrogel treated wound area was effectively reduced by about 60%, indicating that the PNIPAm-P hydrogel had excellent temperature-triggered contractile behavior, making it possible to mechanically promote wound closure.

Example 7: wound healing study of PNIPAm-P hydrogel on healthy BALB/c mice

The back of the mice was first shaved and disinfected with iodophors and 75% alcohol, and then two wounds (one on each side) were created on the back of the mice using a dermatome (diameter: 6 mm). The acrylate splint was then secured to the wound edge of the mouse. The PNIPAm-P hydrogel prepared in example 1 was placed on a skin wound of a healthy mouse. Other materials and medical bandages were then used to cover the wound area to minimize moisture loss and prevent scratching by the animal. Wounds were measured and analyzed every two days by observation with Image J software. The results are shown in FIG. 6.

Figure 6 is a graph of wound healing after dressing treatment. Fig. 6(a) is a representative image of the wound on

days

0, 2, 4 and 6 of treatment, and fig. 6(b) is the quantitative statistics of fig. 6 (a). As shown in the figure, PNIPAm-P showed better wound closure effect at the earliest day 2 and reached at day 6

The closing rate of (A) is far better than that of the control group

PAAm-P group

And PNIPAm group

The PNIPAm-P is shown to have obvious effect on wound treatment of healthy mice.

Example 8: macrophage changes in wound tissue following PNIPAm-P treatment

In a wound model of mice, mice were sacrificed and their wound tissues were collected to study changes in wound microenvironment on day 5 after different treatments. Freshly collected wound tissue was digested with collagenase (1mg/mL) and hyaluronidase (1mg/mL) at 37 ℃ for 30min and centrifuged at 14800rpm for 10min to remove the supernatant. Then, single cells were prepared by passing through a single cell filter membrane at a ratio of 0.2. mu.L/10⁶Individual cell ratios were stained with flow antibody for 1h at 37 ℃. Finally, the antibody was washed away and detected by flow cytometry. The results are shown in FIG. 7.

FIG. 7 is a graph of macrophage changes at the wound site. Fig. 7(a) and 7(b) are representative images and quantitative statistics of flow cytometry analysis of macrophages (CD11b + F4/80+), and fig. 7(c) and 7 (d) are representative images and quantitative statistics of flow cytometry analysis of M2-class macrophages (CD206+ CD11b + F4/80 +).

As shown, the total number of macrophages remained nearly constant in the wounds treated with the different hydrogel dressings, while the number of M2 phenotype macrophages was significantly increased around the wounds treated with the PNIPAm-P hydrogel dressing. This indicates that PMAA in PNIPAm-P is effective in promoting the polarization of macrophages from a "pro-inflammatory" phenotype (type M1) to an "anti-inflammatory" phenotype (type M2).

Example 9: inflammatory conditions of wound tissue following PNIPAm-P treatment

Mice were sacrificed and their wound tissue was collected, freshly collected wound tissue was minced and incubated with 500 μ L of cell lysate for 30min at 4 ℃, followed by centrifugation at 14800rpm for 10min to remove the pellet. The concentration of major inflammation-associated cytokines in the wound tissue supernatant was measured by enzyme-linked immunosorbent assay (ELISA). The results are shown in FIG. 8.

FIG. 8 is a graph showing the expression of inflammation-associated cytokines on day 5 of wound treatment. As shown, various proinflammatory cytokines including TNF- α and IL-6 were significantly down-regulated in PNIPAm-P treated wounds at slightly higher concentrations than in PAAm-P treated wounds. This is probably due to the smaller pore size of the PNIPAm-P hydrogel at body temperature limiting the PMAA exposure. The PNIPAm-P can effectively relieve the inflammatory reaction at the wound.

Example 10: vascular condition of wound tissue after PNIPAm-P treatment

On day 15 of wound treatment, mice were anesthetized and coated with a coupling agent at the wound site, followed by PA imaging of neonatal wound tissue using the Oxy-Hemo modality on the Vevo LAZR system. The total percentage of average sO2 in regenerated tissue was quantified using the Vevo software tool. The results are shown in FIG. 9.

Figure 9 is a graph of angiogenesis after dressing treatment. Fig. 9(a) is a representative photoacoustic image in neonatal wound tissue, and fig. 9(b) is a quantitative statistic of oxyhemoglobin saturation. As shown, wounds treated with PNIPAm-P hydrogel showed significantly higher mean blood oxygen levels. The PMAA in the PNIPAm-P hydrogel is shown to be capable of effectively promoting the generation of new blood vessels at the wound part.

Example 11: wound healing study of PNIPAm-P hydrogel on healthy Bama miniature pigs

The backs of the piglets were first shaved and disinfected with iodophors and 75% alcohol, then eight wounds of local thickness (750 μm deep, 2.5 x 2.5cm) (four on each side) were created on the backs of the piglets with a skin knife, the minimum distance between the wounds being kept at 4 cm. The PNIPAm-P hydrogel prepared in example 1 was then applied to the right side wound and the left side wound was used as a control. The wound was then covered with a layer of transparent dressing and tacked to the skin. For further protection, a layer of flexible self-adhesive bandage is used to support and protect the underlying dressing. After the experiment, the piglets were monitored for any discomfort and observed every two days by Image J software, the wounds were measured and analyzed. The results are shown in FIG. 10.

Figure 10 is a graph of wound healing after dressing treatment. Fig. 10(a) is a representative image of the wound on

days

0, 2, 4, 6 and 7 of treatment, and fig. 10(b) is a quantitative statistic of fig. 10 (a). As shown, the wounds treated with PNIPAm-P showed an accelerated wound healing process, with the PNIPAm-P treatment wound healing rate approaching 60% at day 7, while the control wound reached only 20% healing rate. The PNIPAm-P is proved to have obvious effect on treating the wounds of the healthy miniature pigs.

Example 12: study of PNIPAm-P treatment of diabetic wounds

A diabetic mouse model was induced by intraperitoneal injection of Streptozotocin (STZ) into healthy C57BL/6 mice according to standard protocols. The fasting blood glucose level in C57BL/6 mice was more than 11.1mmol/L for three consecutive days after STZ injection (50 mg/kg per mouse for 5 consecutive days), indicating that a diabetic mouse model was successfully established. Wound modeling and treatment procedures were performed according to the method of example 7. Wounds were observed and recorded every two days, and measured and analyzed by Image J software. The results are shown in FIG. 11.

Figure 11 is a graph of wound healing after dressing treatment. Fig. 11(a) is a representative image of the wound on

days

0, 2, 4 and 6 of treatment, and fig. 11(b) is the quantitative statistics of fig. 11 (a). As shown, significant improvement in diabetic wound closure was observed at the earliest day 2 in the PNIPAm-P treatment group. It reached-45% closure rate on day 2, which was much higher than the control group

PAAm-P group

And PNIPAm group

The PNIPAm-P has obvious effect on treating the wounds of the diabetic mice.

Example 13: potential repair effect of PNIPAm-P on in-vitro organs

New Zealand laboratory rabbits were sacrificed to collect their major organs, including heart, liver, kidney and stomach. Tissue defects (8mm) were made on each organ using a dermatome and covered with PNIPAm-P and pressed for 10 min. These organs were then incubated at 37 ℃ for 2 hours. Finally the dressing was peeled off and the wound area was recorded and calculated by Image J software. The results are shown in FIG. 12.

FIG. 12 is a graph showing the effect of PNIPAm-P contraction on a defective region of an organ in vitro. As shown, PNIPAm-P is indeed effective in promoting the contraction of defective areas of different organs, in particular the liver. This suggests that PNIPAm-P with active contractile behavior has the potential for tissue repair in vivo.

The above-mentioned embodiments are merely preferred embodiments for fully illustrating the present invention, and the scope of the present invention is not limited thereto. The equivalent substitution or change made by the technical personnel in the technical field on the basis of the invention is all within the protection scope of the invention. The protection scope of the invention is subject to the claims.

Claims

1. The hydrogel dressing is a semi-interpenetrating network hydrogel obtained by performing free radical polymerization on poly (N-isopropylacrylamide), polymethacrylic acid and gelatin.

2. The hydrogel dressing according to claim 1, wherein the hydrogel dressing is synthesized from poly (N-isopropylacrylamide), polymethacrylic acid and gelatin in a mass ratio of 1: 1: 1-4: 1: 1.

3. the hydrogel dressing of claim 1, wherein the hydrogel dressing has pore sizes of 7 to 8 μm at 25 ℃; the pore diameter of the hydrogel dressing is 4-5 mu m at 37 ℃.

4. A method of manufacturing a hydrogel dressing according to any one of claims 1 to 3, comprising the steps of:

5. The method of claim 4, wherein the polymethacrylic acid has a molecular weight of 6-100 kDa.

6. The method of claim 4, wherein the cross-linking agent comprises N, N' -methylenebisacrylamide, ethylene glycol diacrylate, m-phenylenedimaleimide, pentaerythritol triacrylate.

7. The method of claim 4, wherein the initiator comprises persulfate, hydrogen peroxide, and water-soluble azo initiator, and the accelerator comprises tetramethylethylenediamine and triethylamine.

8. The method of claim 4, wherein the solvent used to prepare the sodium polymethacrylate, the gelatin solution and the poly (N-isopropylacrylamide) solution is ultrapure water.

9. The method of claim 4, wherein the method further comprises the steps of immersing the hydrogel in saline and lyophilizing the hydrogel dressing after gelation.